A stem cell is a cell type that has a unique capacity to renew itself and to give rise to specialized or differentiated cells. Although most cells of the body, such as heart cells or skin cells, are committed to conduct a specific function, a stem cell is uncommitted, until it receives a signal to develop into a specialized cell type. What makes the stem cells unique is their proliferative capacity, combined with their ability to become specialized. For years, researchers have focused on finding ways to use stem cells to replace cells and tissues that are damaged or diseased. So far, most research has focused on two types of stem cells, embryonic and somatic stem cells. Embryonic stem cells are derived from the preimplanted fertilized oocyte, i.e. blastocyst, whereas the somatic stem cells are present in the adult organism, e.g. within the bone marrow, epidermis and intestine. Pluripotency tests have shown that whereas the embryonic or blastocyst-derived stern cells (hereafter referred to as blastocyst-derived stem cells or (BS cells) can give rise to all cells in the organism, including the germ cells, somatic stem cells have a more limited repertoire in descendent cell types.
In 1998, investigators were for the first time able to isolate hBS cells from human fertilized oocytes and to grow them in culture see e.g. U.S. Pat. No. 5,843,780 and in U.S. Pat. No. 6,200,806.
The procedure used in the patent specifications mentioned above depends on the use of blastocysts with an intact zona pellucida. Furthermore, the method disclosed in these patents specifically use inner cell mass cells that have been isolated by iminunosurgery for plating on mouse embryonic feeder cells. This method has several drawbacks, for example, it is time consuming, technically difficult and results in low yields of stem cells. Taken together, these drawbacks make it a costly method.
The so far few publications in the field illustrate the problems associated with establishing these stein cells from human blastocysts. As a result very few hBS cell lines are available.
Perhaps the most far-reaching potential application of hBS cells is the generation of cells and tissue that could be used for so-called cell therapies. Many diseases and disorders result from disruption of cellular function or destruction of tissues of the body. Today, donated organs and tissues are often used to replace ailing or destroyed tissue. Unfortunately, the number of people suffering from disorders suitable for treatment by these methods far outstrips the number of organs available for transplantation. The availability of hBS cells and the intense research on developing efficient methods for guiding these cells towards different cell fates, e.g. insulin-producing β-cells, cardiomnyocytes, and dopamine-producing neurons, holds growing promise for future applications in cell-based treatment of degenerative diseases, such as diabetes, myocardial infarction and Parldnson's.
A significant challenge to the use of pluripotent stem cells for therapy is that they are traditionally cultured on a layer of feeder cells to prevent differentiation and to promote cell survival and proliferation. Without feeder cells in the culture environment, the stein cells will die, or differentiate into a heterogeneous population of committed cells. Unfortunately, using feeder cells increases production costs, impairs scale-up, and produces mixed cell populations that require the pluripotent stem cells to be separated from feeder cell components. Furthermore, for therapeutic applications it will be of greatest importance that the hBS cells are cultured without xenogenic tissue contact, such as, e.g. feeder cells. Thus, there is a need for developing methods for propagating human blastocyst-derived stem cell lines without the use of feeder cells.
Other potential applications of hBS cells themselves and cell populations derived there from are found e.g. in the drug discovery process in the pharmaceutical industry and in toxicity testings of all kinds of chemicals. Today, large-scale and high throughput screening of drug candidates usually relies on biochemical assays that provide information on compound binding affinity and specificity, but little or no information on function. Functional screening relies upon cell-based screens and usually uses organisms of poor clinical relevance such as bacteria or yeasts that can be produced cheaply and quickly at high volume. Successive rounds of screening use model species of greater clinical relevance, but these are more costly and the screening process is time consuming. Screening tools based on human primary cells or immortalised cell types exist, but these cells are limited in supply or usefulness due to loss of vital functions as a result of in vitro culture and transformation. The access to undifferentiated hBS cells and hBS cells differentiated under engineered conditions and in the absence of interfering feeder cells provides a new and unique capability to conduct human cell-based assays with high capacity, but without compromising clinical relevance.
The following definitions and abbreviations are used herein